This project describes a 5-year career development program designed to foster an academic career in hematologic research and hematopathology. The proposed research program will build on an existing fund of knowledge using the unique resources available at Washington University, which has a proven track record of developing physician scientists. Dr. Timothy Ley, an expert in myeloid development and cancer genomics and a previous recipient of the American Society of Hematology Mentor Award, will serve as the research mentor. The goals of this proposal are to obtain practical and/or didactic training in scientific methodology, technical skills, grant writing, and scientific publication for the purposeof establishing an independent research program focused on hematopoietic development. The proposed research will focus on the Notch pathway to determine the mechanism(s) by which Notch signaling alters myeloid development. Our recent studies have shown that myeloid progenitors expressing the PML-RARA oncogene (using the Ctsg-PML-RARA mouse model) have enhanced self-renewal, which can be abrogated in vitro by Notch inhibition. These observations will be extended in vivo using an established mouse model of Notch inactivation (conditional DN-MAML-GFP) in the following Aims: Specific Aim 1: We will investigate the role of Notch signaling in self-renewal and myeloid development in vivo. The impact of Notch blockade in both normal and Ctsg-PML-RARA-derived hematopoietic stem/progenitor cells (HSPCs) will be assessed by in vivo methods to assess progenitor fitness and signaling, as well as the long-term development of myeloid neoplasms; and Specific Aim 2: We will identify Notch target genes and epigenetic changes in early myeloid progenitors. The GFP moiety on the DN-MAML1-GFP fusion will be targeted for chromatin immunoprecipitation followed by Illumina paired-end sequencing (ChIP-Seq) to determine the genomic targets of Notch activation in c-Kit positive cells (enriched in myeloid stem/progenitors). The studies will be paired with transcriptional profiling and additional ChIP-Seq experiments to monitor histone modifications. These studies will define the mechanisms by which the Notch pathway influences myeloid development and will identify myeloid-specific Notch functions, which may ultimately guide novel therapeutic approaches for myeloid diseases.